The HIV and HSV-2 syndemic is well recognized, but the biological mechanisms that contribute are not understood. The recent recognition that HSV-2 is characterized by a frequent state of subclinical shedding suggests that the virus might contribute to persistent immune activation and prompted us to examine the impact of HSV-2 on peripheral blood T cells and on HIV reservoirs. Taking advantage of our biorepository of peripheral blood mononuclear cells (PBMC) from well-characterized HIV+ women on antiretroviral therapy who were or were not HSV-2 seropositive (HIV+/HSV-2+ vs. HIV+/HSV-2-), we found a significant difference in the phenotype of CD4+ (but not CD8+) T cells in HIV+/HSV-2+ compared to HIV+/HSV-2- women. These changes included an increase in the frequency of activated cells, but a paradoxical decrease in the expression of IL-32, an intracellular cytokine presumed to be associated with inflammation. Moreover, when CD4+ T cells isolated from virally suppressed HIV+/HSV-2+ women were stimulated with latency reversal agents, the addition of recombinant IL-32 to the cultures blocked HIV reactivation. These observations suggest that IL-32 plays a pivotal role in controlling HIV reactivation and suggest a new paradigm underlying the HIV-HSV-2 syndemic. We hypothesize that HSV-2 triggers changes in local (at the site of HSV-2 genital skin outbreaks) and peripheral blood CD4+ T cells including a reduction in intracellular IL-32 levels that promote HIV reactivation. Conversely, high levels of IL-32 contribute to the maintenance of HIV reservoirs suggesting that IL-32 blockade may synergize with strategies to reactivate HIV as part of a ?shock and kill? approach to cure. To test these hypotheses, we will analyze serial samples of PBMC from HIV infected women before and after HSV-2 acquisition from two unique cohorts: women enrolled in Microbicides Trial Network (MTN)-015, a longitudinal study of African women who seroconverted to HIV while participating in pre-exposure prophylaxis trials, and U.S. women with established HIV infection enrolled in the Bronx Women's Interagency Study (WIHS). We will also compare PBMC in HIV-infected men who are or are not coninfected with HSV-2. We will phenotype immune cell subpopulations to define the changes that occur in association with HSV-2 acquisition and the impact of these changes on plasma viral loads and HIV reservoirs. We will prepare CD4+ T cell libraries of IL- 32lo cells and determine whether these subpopulations are enriched in HSV-2 and/or HIV reactive cells and whether decreased IL-32 interferes with immune functions. We will also take advantage of our repository of genital skin biopsies (herpes lesion and unaffected contralateral side) and analyze the CD4+ T cells to determine whether they are enriched for cells of specific phenotypes in situ. We will determine how IL-32 blocks the response to latency reactivating stimuli and how IL-32 antagonists promote HIV reactivation. These studies will identify pathways and molecules that could be targeted to block HIV reactivation in response to HSV-2 or conversely, enhance latency reversal as part of ?shock and kill? HIV eradication strategies.